Anticoagulants serve a need in the marketplace in treatment or prevention of undesired thrombosis in patients with a tendency to form blood clots, such as, for example, those patients having clotting disorders, confined to periods of immobility or undergoing medical surgeries. One of the major limitations of anticoagulant therapy, however, is the bleeding risk associated with treatment, and limitations on the ability to rapidly reverse the anticoagulant activity in case of overdosing or if an urgent surgical procedure is required. Thus, specific and effective antidotes to all forms of anticoagulant therapy are highly desirable. For safety considerations, it is also advantageous to have an anticoagulant-antidote pair in the development of new anticoagulant drugs.
Previously reported modified derivatives of factor Xa (fXa), such as those described in U.S. Pat. Nos. 8,153,390 and 8,268,783 (which is herein incorporated by reference in its entirety), including r-Antidote, are useful as antidotes to anticoagulants targeting fXa. The modified derivatives of fXa bind to and/or substantially neutralize the anticoagulant. Certain modifications introduced to fXa, however, pose several challenges for purification since routine methods for purification of clotting factors cannot be used for r-Antidote.